Figure 8

Characteristics of 4T1, 67NR, and E0771 mouse mammary gland cell lines on calcitriol treatment in vitro in the context of inflammatory factors expression. (A) Real-time PCR analysis of Il23, Il6, and Fn1 gene expressions in 4T1 cells treated with 100 nM calcitriol for 72 h (4T1 cal) in vitro. Untreated cells served as the calibrator, and Hprt was used as an endogenous control, N = 3 or 4. (B) IL-6 concentration using ELISA in culture media from 4T1 cells treated with 100 nM of calcitriol for 72 h and then additionally treated with FBS free medium for 24 h. (C) Western blot analysis of cyclooxygenase-2 (COX-2) in 4T1, 67NR, and E0771 cells treated with 100 nM of calcitriol for 72 h. (D) Comparison of COX-2 expression between the three mouse mammary gland cancer cell lines (western blot). (C) and (D) Next to the graphs, representative blots are presented. (E) Prostaglandin E2 (PGE2) concentration using ELISA in culture media from 4T1 cells treated with 100 nM of calcitriol for 72 h and then additionally treated with FBS-free medium for 24 h. Statistical analysis: (A) Unpaired t-test; (B), (D) and (E) Sidak’s multiple comparisons test; (C) Dunn's multiple comparisons test; *P < 0.05, **P < 0.01.